Despite considerable efforts to develop new therapies for melanoma, patients with advanced disease continue to have a poor prognosis. We have pursued an alternate strategy to develop safe and effective preventive agents, focusing on patients with high familial risk for melanoma. Individuals with a mutant CDKN2A (p16) allele have an 80% lifetime-risk of developing melanoma. We have recruited large numbers of melanoma-prone individuals and used patient-derived tissues to initiate investigation of molecular markers and pathways of melanoma predisposition. Preliminary transcriptomal (microarray) analyses of tissues from one high-risk kindred with an inherited p16 mutation revealed dysregulation of oxidative stress pathways in mutation carriers. We hypothesize that cells with p16 mutations have alterations in oxidative stress responses and that oxidative damage pathways can be targeted for the development of novel chemopreventive agents for melanoma. Our long-term goal is to engage our high-risk cohort in a melanoma chemoprevention trial using an oral antioxidant such as N-acetyl cysteine (NAC). Potential agents can be screened in mouse models of high-risk melanoma. As we continue to recruit patients for an eventual trial, we propose here the following specific aims: 1) validate oxidative damage and dysregulation of oxidative stress responses in p16-mutant skin tissues; 2) assess ultraviolet (UV)-induced oxidative damage and responses in normal human melanocytes, cells with RNAi-mediated p16 deficiency and nevomelanocytes grown from p16-mutant nevi, and protection by NAC; and 3) test the capacity of NAC to prevent UV-induced melanoma in melanoma-prone mice. It should be recognized that the same susceptibility genes identified in familial studies undergo somatic mutations in the pathogenesis of sporadic melanoma. Thus, agents that effectively reduce risk in familial patients are also likely to be useful in the prevention of sporadic melanoma. [unreadable] Relevance to public health: Our studies in patients with inherited risk of melanoma have provided a clue that may eventually enable us to prevent this disease. Here, we propose to examine mechanisms in patient- derived cells and the ability of an antioxidant to prevent melanoma in a mouse model. These studies will lead to formal testing whether an oral antioxidant can prevent melanoma in patients with increased risk. [unreadable] [unreadable] [unreadable]